The molecular basis for the cariogenicity of Streptococcus mutans will be further investigated utilizing molecular genetic techniques. Since the conversion of sucrose to insoluble glucan and lactic acid play key roles in this process, the genes involved will be examined. The gene coding for mutansynthetase activity will be isolated with a phage cloning system and investigated along with the previously isolated dextransucrase gene. Both genes will be sequenced, their regulatory regions identified and compared for homology. In addition, in vitro mutagenesis of the genes will be carried out to identify regions of the proteins required for enzymatic activity. These altered genes will also be reintroduced into strain GS-5 to produce specific lesions in the organism in order to examine the respective role of each enzyme in cariogenicity. The previously cloned fructosyltransferase gene will also be subjected to in vitro mutagenesis followed by transformation into strain GS-5 in order to determine the number of fructosyltransferase genes in the organism as well as the role of the cloned gene in pathogenicity. The gene will also be sequenced in order to investigate the secretion of the enzyme and its enzymatic properties. The previously cloned sucrose-6-phosphate hydrolase gene will be sequenced in order to determine the genetic basis for its strong expression in Escherichia coli. In vitro generated mutants of this gene will be utilized to clone the sucrose enzyme II of the sucrose PT system in order to investigate the genetic basis for sucrose transport in S. mutans.